Objective To investigate the effect of CaMK Ⅱ expression on apoptosis of rat hepatocytes BRL-3A.Methods Rat BRL-3A cells were stable passage were cultured.The CaMK Ⅱ γ protein (LV-CaMK Ⅱ γ group) and CaMK Ⅱ γshRNA (shRNA group) lentiviral expression systems were constructed.The corresponding blank vectors (LV-NC group and shRNA-NC group) and normal saline (CON group) were perfused into the control groups.The expression levels of CaMK Ⅱ,Cyt C and MF proteins were detected by Western blotting,and the apoptosis rate of BRL-3A cells was measured by Tunel method.Results The protein expression of CaMK Ⅱ,Cyt C and AIF in LV-CaMK Ⅱ γ group was significantly higher than that in CON group (P＜0.05).The protein expression of CaMK Ⅱ,Cyt C and AIF in shRNA group was significantly lower than that in CON group (P＜ 0.05).There was no significant difference among CON group,LV-NC group and shRNA-NC group (P＞0.05).At the same time point,the apoptosis rate of hepatocytes in LV-CaMK Ⅱ γ group was significantly higher than that in CON group (P＜0.05).At the same time point,the apoptosis rate of hepatocytes in shRNA group was significantly higher than in CON group (P＜0.05).There was no significant difference in the apoptosis of hepatocytes among CON group,LV-NC group and shRNA-NC group (P＞0.05).Conclusion The specific CaMK Ⅱ signaling pathway can inhibit the apoptosis of BRL-3A cells,while the enhanced CaMK Ⅱ signaling pathway promotes the apoptosis of BRL-3A cells.

Objective To observe the effect of Oxycontin combined with Gabapentin for treatment of malignant neuropathic pain.Methods Sixty-three cases of malignant neuropathic pain in Jinshan Hospital Affiliated to Fudan University were randomly divided into group A, B and C.Patients of which were given Oxycontin, Gabapentin, Oxycontin combined with Gabapentin respectively for pain treatment.The analgesic effects, toxic reaction side effects, quality of life, and immune function were all compared in three groups.Results Compared with pretherapy, the cancer pain score (NRS), quality of life (QOL) and karnofsky performance status(KPS) scores in all groups were changed significantly after drugs therapy(F=375.852,154.612, 151.838,P＜0.05).The levels of CD3,CD4, CD4/CD8 and NK cells in all groups were higher than before therapy(F=158.935,108.145,366.973,92.090,P＜0.05).After treatment,the NRS, QOL and KPS scores in group C were 2.00± 0.86,44.80± 6.07, 84.50± 6.05, in group A were 3.35 ± 0.67,37.35 ± 5.71,74.50 ±10.99,and in group B were 4.05±0.94,35.85±5.90,72.00±8.34, and the different were significant (F =3.250,10.499,3.465,P＜0.05).The levels of CD3, CD4, CD4/CD8and NK cells in group C were (72.94 ±5.63)％,(41.52±4.19)％, 1.86±0.30, (27.57±6.86)％,in group A were (62.84±5.27)％, (33.84 ±5.40)％,1.35±0.37, (20.49±6.67) ％,and in group B were (62.22±8.10)％, (33.19±6.90)％, 1.32 ± ±0.41, (20.32±5.63) ％, and the different were significant (F =3.377,3.344,3.352,3.386, P＜ 0.05).The patient in group C had less adverse effects than those in group A and B.Conclusion Oxycontin and Gabapentin in treatment of malignant neuropathic pain is effective.

Objective To explore the relationship between serum HBV DNA level , expression of PD-1 on CTL and liver functions in patients with hepatitis B virus cirrhosis. Methods 109 HBV hepatic cirrhosis patients with , HBV DNA positive , HBeAg positive and HLA-A2 positive were divided into two groups according to their HBV DNA levels,52 cases in group A, HBV DNA 2-4 log10 copies/mL, 57cases in group B, HBV DNA 5-7 log10 copies/mL , differences of HBV specific CTL surface PD-1 expressions , HBV specific CTL levels and liver functions were compared between two groups. Results HBV specific CTL surface PD-1 expressions of group A were lower than that of group B (t = 11.101, P < 0.01), HBV specific CTL levels of group A were higher than that of group B (t = 24.424, P < 0.01), ALT of group A were lower than that of group B (t = 2.652, P < 0.01), ALB of group A were higher than that of group B (t = 2.347, P < 0.05). Child-pugh rating, grade C of group A was lower than that of group B (χ2= 4.262,P < 0.05). Conclusions HBV specific CTL surface PD-1 expression levels were higher in those hepatic cirrhosis patients with , high level of serum HBV DNA, and HBV specific CTL levels were lower, liver functions damage were much serious. So, lower HBV DNA levels , lower HBV specific CTL surface PD-1 expression levels and higher HBV specific CTL levels can product relatively good effect of anti-viral treatment.

Objective To explore the changes of programmed death receptor-1 (PD-1) in chronic hepatitis B (CHB) patients with different baseline of hepatitis B virus (HBV) DNA levels after treatment with adefovir dipivoxil (ADV).Methods One hundred CHB patients with positive hepatitis B e antigen (HBeAg),1 × 104 copy/mL≤HBV DNA≤1 × 107 copy/mL,and positive human leukocyte antigen-A2 were divided into two groups according to the baseline HBV DNA level:47 cases in low virus load group whose HBV DNA level was ≤1 × 105 copy/mL; 53 cases in high virus load group whose HBV DNA level was＞1 × 105 copy/mL.Both groups were treated with ADV 10 mg/d.Serum HBV DNA,HBeAg seroconversion rate,alanine aminotransferase (ALT) and total bilirubin (TBil) levels of both groups before treatment and 12 months after treatment were compared.Flow cytometry was used to test peripheral blood HBV-specific cytotoxic T lymphocyte (CTL) surface PD-1 and peripheral blood HBV-specific CTL level.Categorical data were tested by x2 test; quantitative data was compared with t-test.Results Peripheral blood HBV-specific CTL surface PD-1 of CHB patients in low virus load group was 20.17 ％±1.69％,which was lower than that in high virus load group (41.38％±2.30％,t =53.02,P＜0.01) ; peripheral blood HBV specific CTL levels in two groups were 0.37％±0.02％ and 0.17％± 0.02％,respectively (t=50.47,P＜0.01) ; ALT and TBil levels in low virus load group were both lower than those of high virus load group (t=13.07,P＜0.01; t=5.06,P＜0.01).Twelve months after treatment,HBV DNA of 25 cases (53.2％) in low virus load group and 10 cases (18.9％) in high virus load group were lower than the detectable level (HBV DNA＜500 copy/mL,x2 =12.89,P＜0.01);HBeAg seroconversion was achieved in 15 cases(31.9％) and 1 case (1.9％),respectively (x2 =16.72,P＜0.01) ; peripheral blood HBV-specific CTL surface PD-1 expression levels were 9.00 ％ ±1.38 ％ and 29.40 ％ ± 3.76 ％,respectively (t =36.80,P＜ 0.01) ; peripheral blood HBV-specific CTL levels were 0.65％±0.10％ and0.48％±0.07％,respectively (t=9.61,P＜0.01).Conclusions After treatment with ADV,along with the decrease of HBV DNA load,HBV-specific CTL surface PD-1 expression decreases,while HBV-specific CTL level increases.The changes in low virus load group are much more remarkable.

Objective To investigate the association between the difference of specific cytotoxic lymphocyte (CTL) and liver function of patients with hepatitis B virus (HBV) genotype B and C infections and interleukin (IL)‐7 induced follicular helper T lymphocytes (Tfh) .Methods Sixty‐seven patients with chronic hepatitis B (CHB) hospitalized at Wuxi No .5 People′s Hospital from August 2013 to January 2015 were collected and 30 healthy blood donors were set as healthy control group .The peripheral blood IL‐7 , Tfh ,IL‐21 ,HBV specific‐CTL ,nonspecific CTL ,levels of HBV DNA ,alanine aminotransferase (ALT) and total bilirubin (TBil) were compared between patients with genotype B and C infection ,hepatitis B e antigen (HBeAg)‐positive and HBeAg‐negative CHB ,high ALT level and low ALT level .Multivariate regression analysis was performed to determine the factors associated with IL‐7 .The t test was used for quantitative data and chi‐square test was used for categorical data .Results Of the 67 CHB patients with average age of (35 .1 ± 11 .4) ,48 were male and 19 were female;32 were infected with genotype C and 35 were infected with genotype B ;40 were HBeAg‐positive CHB and 27 were HBeAg‐negative CHB ;17 were with high ALT levels and 50 were with low ALT levels .IL‐7 ,Tfh ,IL‐21 and HBV specific‐CTL levels in the peripheral blood of genotype C‐infected patients were (20 .79 ± 4 .82 ) ng/L , (3 .93 ± 0 .82)% ,(24 .77 ± 7 .52) ng/L and (0 .20 ± 0 .04)% ,respectively ,while in genotype B‐infected patients , those were (29 .13 ± 8 .17) ng/L ,(5 .92 ± 1 .92)% ,(39 .94 ± 24 .00) ng/L and (0 .40 ± 0 .06)% , respectively .Levels of IL‐7 , Tfh ,IL‐21 and HBV specific‐CTL in genotype C‐infected patients were significantly lower than those in genotype B‐infected patients (t= 5 .027 ,5 .595 ,3 .553 and 15 .133 , respectively ;all P<0 .01) .Nonspecific CTL ,HBV DNA ,ALT and TBil levels in the peripheral blood of genotype C‐infected patients were all significantly higher than those in genotype B infected‐patients (t=4 .899 ,6 .815 ,2 .763 and 4 .899 ,respectively ;all P< 0 .01) .IL‐7 ,Tfh ,IL‐21 ,HBV specific‐CTL levels in the peripheral blood of HBeAg‐positive patients were significantly lower than those in HBeAg‐negative patients (all P<0 .01) .Nonspecific CTL ,HBV DNA ,ALT and TBil levels in the peripheral blood of HBeAg‐positive patients were all significantly higher than those in HBeAg‐negative patients (all P<0 .05) .IL‐7 ,Tfh ,IL‐21 ,HBV specific‐CTL levels in the peripheral blood of patients with high ALT levels were all significantly lower than those in patients with low ALT levels (all P<0 .01) .Nonspecific CTL and HBV DNA levels in the peripheral blood of patients with high ALT levels were both significantly higher than those in patients with low ALT levels (both P<0 .01) .HBV DNA ,IL‐21 and nonspecific CTL were all correlated with IL‐7 (all P<0 .01) .Conclusion The differences of HBV specific‐CTL and liver function in CHB patients infected with genotype B and C may be correlated with interleukin‐7 induced Tfhcells.

Abstract BACKGROUND: Looking for the early diagnosis of acute rejection indicators after liver transplantation can assess the risk after liver transplantation quickly and effectively, and T lymphocytes play the significant role in acute rejection. OBJECTIVE:To observe the relationship between acute rejection and variation of expression of T cel subset in blood after liver transplantation in rhesus monkey. METHODS: The sixteen liver transplant models in rhesus monkey which were constructed successfuly by the method of “double-cuff and one support tube” were divided into two groups randomly: experiment group (no treated by immunosuppressant in perioperative period) and control group (treated by immunosuppressant in perioperative period). Then the blood specimen and liver tissue respectively were colected at 6, 12, 24 and 72 hours after operation. The levels of alanine transferase, aspartate aminotransferase, and total bilirubin were detected with the fuly automatic biochemical analyser. The levels of CD4+/CD8+were tested by flow cytometry. The liver tissue in rhesus monkey after liver transplantation was detected by hematoxylin-eosin staining. The degree of acute rejection was evaluated by Banff Score System. RESULTS AND CONCLUSION: Acute rejection appeared in the experiment group at 12, 24, and 72 hours after liver transplantation. Levels of alanine transferase, aspartate aminotransferase, and total bilirubin were significantly higher in the experimental group than in the control group at 24 and 72 hours after transplantation (P < 0.05). The expression of CD4+/CD8+of the experiment group and control group began to rise at 6 hours after surgery, but the experiment group increased the most obvious. CD4+/CD8+ expression was significantly greater in the experimental group than in the control group at 24 and 72 hours after transplantation (P < 0.05). Morphological pathology was severer, and Banff score was higher in the experiment group than in the control group at 72 hours (P < 0.05). These data suggested that the variation of expression of CD4+/CD8+was earlier than the change of liver tissue pathology and the change of liver function in the early acute rejection after liver transplantation. The rise of level of CD4+/CD8+ after liver transplantation indicated the increase of celular immunity in body, which had an important role in the early diagnosis of acute rejection after liver transplantation.

Objective To explore the effect of nano-hydroxyapatite/collagen scaffolds incorporating ADM-PLGA microspheres in repairing large bone defects of rabbit femoral condyle. Methods Animal models of bone defects were induced in 21 New Zealand white rabbits by drilling holes in bilateral femoral lateral condyles , and the rabbits were equally divided into 3 groups:group A as the control group with the defects untreated , group B treated by filling with nano-hydroxyapatite/collagen scaffolds (NHAC), and group C treated by filling with the nano-hydroxyapatite/collagen scaffolds incorporating ADM-PLGA microspheres (ADM-PLGA-NHAC). At week 12 after implanting , the rabbits were all sacrificed for the implanted scaffolds , which were then examined by X-ray , and Micro-CT 3D reconstruction and in histology for evaluation of the new bone formation. Results X-ray, Micro-CT and the measurement and analysis of BMD indicated thatthere was no significant differencein the new bone formation between group B and group C (P > 0.05). The histological examination revealed that. 12 weeks after operation an evident number of new born bones were seen on the implanted scaffolds in groups B and C , while very few were seen scattering in group A. Conclusion The nano-hydroxyapatite/collagen scaffolds incorporating ADM-PLGA microspheres is effective in repairing bone defect without influencing the prosthetic process.

Objective To evaluate the effect of comprehensive intervention on perioperative antimicrobial prophy-laxis in clean incision surgery in a hospital.Methods From 2011 ,clean incision surgery cases were performed com-prehensive intervention,antimicrobial use in 2011 -2013 were compared.Results A total of 5 945 cases of clean in-cision surgeries were investigated between 2011 and 2013,3 827 cases (64.37%)received prophylactic use of anti-microbial agents.Prophylactic antimicrobial usage rates in 2011 -2013 were 84.95%,69.99%,and 52.97% re-spectively(χ2 =380.94,P <0.001);the correct rates of medication time were 50.97%,79.99%,and 98.95% re-spectively(χ2 =827.02,P <0.001 );the percentages of prophylactic antimicrobial use ≤24 hours were 24.91 %, 39.96%,and 64.95% respectively(χ2 =422.55,P <0.001 );additional antimicrobial usage rates during surgery were 50.00%,60.00%,and 80.00% respectively(χ2 =59.47,P <0.001 ).Conclusion The implementation of comprehensive intervention measures can standardize antimicrobial use,reduce prophylactic antimicrobial usage rate,improve the correct rate of medication time,shorten the duration of antimicrobial use,and implement addition-al use of antimicrobial agents during surgery.

BACKGROUND:Interleukin-6 is an important cytokine in the immune inflammatory response, strongly links with graft rejection reaction, and plays an important role in diagnosis of graft rejection and evaluation of anti-rejection. OBJECTIVE:To measure the expression of interleukin-6 in acute rejection of the liver transplantation in the rhesus monkey, and to evaluate the value as an early diagnosis of acute rejection after liver transplantation. METHODS:A total of 16 rhesus monkeys were used as the object and randomly divided into experimental group (no treated by immunosuppressant in perioperative period), and control group (treated by immunosuppressant in perioperative period). The al ograft orthotopic liver transplantation models were established in those monkeys. Then serum and liver tissue were col ected at 6, 12, 24, and 72 hours after surgery. Al ograft rejection was monitored by liver function tests, and hematoxylin-eosin staining of liver and Banff score. Final y, the expression levels of interleukin-6 were detected by enzyme linked immunosorbent assay and immunohistochemistry.RESULTS AND CONCLUSION:Acute graft rejection reaction appeared at 12, 24 and 72 hours after liver transplantation in the experimental group. The expressions of alanine aminotransferase, aspartate aminotransferase and total bilirubin were significantly higher in the experimental group than in the control group at 24 and 72 hours (P<0.05). Histological manifestations were severer and Banff score was higher in the experimental group at 72 hours than in the control group (P<0.05). Interleukin-6 levels were significantly higher in the serum and liver tissue of experimental group than in the control group at 12, 24 and 72 hours after liver transplantation (P<0.05), especial y at 72 hours. Results suggested that interleukin-6 possibly participated in rejection after liver transplantation. The expression of interleukin-6 was probably of significance in the early diagnosis of acute rejection after orthotopic liver transplantation in rhesus monkeys.

BACKGROUND:Tumor necrosis factor-αis an inflammatory cytokine involved in the immune response and increasing graft antigen expression. OBJECTIVE:To investigate the relationship between tumor necrosis factor-αin the liver tissue and acute rejection after liver transplantation in a rhesus monkey. METHODS:Liver transplant models in rhesus monkey were constructed by the improved vascular dual cuff, supporting tube of biliary tract and artery anastomosis method. The successful models were randomly divided into experimental group (no immunosuppressant treatment in perioperative period) and control group (treated by immunosuppressant in perioperative period). Then the blood samples and liver tissue were col ected at 6, 12, 24, and 72 hours after surgery. Al ograft rejections of liver transplantation were monitored by liver function tests, hematoxylin-eosin staining and Banff score. Final y, the expression level of tumor necrosis factor-αwas detected by western blot analysis and immunohistochemistry technique. RESULTS AND CONCLUSION:The expression of tumor necrosis factor-αin the experimental group and control group began to increase at 6 hours, reached the peak at 12 hours, and then decreased at 24-72 hours. The changes of expression level were the most obvious in the experimental group. At 6, 12, 24 and 72 hours, the expression of tumor necrosis factor-αin the experimental group was significantly higher than that in the control group (P<0.05). This change appeared earlier than pathological changes in the liver and liver function. Variations in the expression of tumor necrosis factor-αafter liver transplantation have important implications for early diagnosis of acute rejection after liver transplantation.